Exposure of cells, tissues and organs to “stress,” such as elevated temperature, heavy metals, oxidants, chemicals interfering with mitochondrial function, alcohols, hypoxia, hyperosmotic and hypoosmotic environments, amino acid analogues, and benzoquinone ansamycins results in the activation or enhanced activity of a group of genes known as heat shock or stress (hsp) genes. See, for example, Scharf et al., “Heat Stress Promoters and Transcription Factors,” in Results and Problems in Cell Differentiation 20, Nover (Ed.), pages 125-162 (Springer-Verlag 1994). When cells, tissues and organs are returned to a normal condition, stress gene activity declines, until it reaches the low pre-stress level.
Stress genes encode a small number of heat shock or stress protein (Hsp) families. Major families of stress proteins are distinguished on the basis of molecular weight and amino acid sequence. See, for example, Nover and Scharf, Cell. Mol. Life Sci. 53:80 (1997). They include Hsp 110 (Hsp's with a subunit molecular weight of about 110 kDa), Hsp 104, Hsp90, Hsp70, Hsp60, Hsp27, Hsp10 and ubiquitin. Many of these Hsp's are molecular chaperones that participate in such basic cellular processes as protein folding, protein degradation and protein trafficking.
Promoter regions of stress genes typically include so-called heat shock element (HSE) sequences. These sequences are essential to render the genes activatable by stress. HSE provide binding sites of proteins named heat shock transcription factors (HSF). See, for example, Wu, “Heat Shock Transcription Factors: Structure and Regulation,” in Annu. Rev. Cell Dev. Biol. 11:441 (1995); Nover and Scharf, Cell. Mol. Life Sci. 53:80 (1997). Mammalian cells can express at least three different HSF. It is thought that the factor termed HSF1 is responsible for the regulation of hsp genes by stress. HSF1 is continuously and ubiquitously expressed in mammalian cells. In the absence of stress, the factor is present in an inactive form, unable to bind HSE sequences of stress gene promoters and to enhance their transcription. During stress, HSF1 is activated, and in the activated form, binds HSE DNA and stimulates transcription of stress genes. Subsequent to a stressful event, the factor relatively rapidly returns to its inactive form. Consequently, transcription of stress genes ceases. Mutant forms of HSF1 have been constructed that are capable of constitutively transactivating stress genes, in the absence of stress.
Promoters of stress genes have been linked to genes of interest to render the genes activatable by stress. Constructs of this kind were used to prepare cell lines, in which genes of interest could be activated by heat or some other form of stress. For example, cell lines have been prepared that contain a stress gene promoter-controlled growth hormone gene. Dreano et al., Gene 49:1 (1986). Moreover, transgenic flies and transgenic nematodes have been produced that express a β-galactosidase gene under stress control. See, for example, Voellmy and Ananthan, U.S. Pat. No. 5,346,812, Candido and Jones, Trends Biotechnol. 14:125 (1996), and Jones et al., Toxicology 109:119 (1996).
A major drawback of the use of stress gene promoters, also referred to herein as stress promoters, to control regulation of a gene of interest is that gene expression induced by a stress promoter can be maintained beyond the duration of the stress treatment only under conditions of extreme stress. Yet such extreme conditions are incompatible with cell survival.
Therefore, a need exists for a means to take advantage of stress-induced gene regulation under conditions that do not endanger cell viability.